E DUARD B U C H N E R Cell-free fermentation Nobel Lecture, December 11, 1907 I would first ask the assembled company to allow me to express my sincere gratitude for having been so highly honoured with the distinction of speak- ing today before the Royal Swedish Academy of Sciences, to which at one time a Scheele and a Berzelius belonged and which at the present time counts Arrhenius among its members, all men whose achievements fill every chem- ist with admiration. The work on which I have to report lies on the bounda- ry between animate and inanimate nature. I therefore have reason to hope that I can interest not only the chemists but also the wide circles of all those who follow the advance of biological science with close attention. It is diffi- cult, however, for a person to be comprehensible and at the same time re- main scientific, so I must ask you to bear with me. If fruit juices or sugar solutions are left to stand in the open air, they show after a few days the processes which are covered by the name of fermenta- tion phenomena. Gas is seen to develop, the clear solution becomes cloudy and a deposit appears which is called yeast. At the same time the sweet taste disappears and the liquid acquires an intoxicating effect. These observations are as old as the hills ; at any rate, these processes have been used since the most ancient times of the human race for the production offermented liquors. It is, however, only since the end of the eighteenth century - i.e. since La- voisier - that we have known that during such a process sugar decomposes into carbon dioxide and ethyl alcohol. A short time later Gay-Lussac was able to show that the weight of the sugar reappears almost exactly in the sum of the weights of these two products of fermentation. The part played by yeast remained for a long time obscure. It was believed that its appearance was of a secondary nature and it was regarded as an inferior kind of precipitation product. The old name for yeast, "Faex cere- visiae", and the expression which has taken root in our language, "die Hefe des Volkes" (the yeast of the people), meaning the outcasts of the nation, also point to this view. Certainly, as early as 1680, the Dutch naturalist Van Leeuwenhoek, who has been called the father of microscopic observation, had established the fairly regular spherical or elliptical shape of yeast, but he 104 1907 E. BUCHNER did not succeed in convincing his colleagues of the vegetable nature of yeast. It was only in the thirties of last century that three researchers, Cagnard Latour in Paris, Theodor Schwann in Berlin and Friedrich Kützing in Nord- hausen, reported almost simultaneously that yeast consisted of live cells of a plant. Though priority in the recognition of the fact belongs to the French- man, who limited himself principally to microscopic observations, Schwann, who also based his work on experiment, was the first to provide the strict proof, and Kützing extended his investigations not only to yeast but also to Theodor Schwann (1810-1882). Founder of cytology, who introduced a new epoch in the study of organisms by establishing that plants and animals are built of elementary components. mother of vinegar, which converts ethyl alcohol into vinegar. The fermen- tation processes thus appeared as a result of the life activity of micro-organ- isms. This vitalistic view, however, received a very mixed reception among the naturalists. In particular there was no lack of keen, even derisive criticism from the greatest chemists of the time, Berzelius, Liebig and Wöhler. Ber- zelius 1 called the new concept of yeast a scientific-poetic fiction. "If the co- alescence of the globules of yeast can be ascribed to the presence of vegetable life, the same reason might well be assumed for the coalescence of globules of clay or calcium phosphate". Liebig and Wöhler 2, however, published a jeering satire in which it was stated that by means of an excellent microscope they had seen the yeast creatures swallow the sugar from the solution, that CELL-FREE FERMENTATION 105 this was instantaneously digested in the stomach and that this digestion could be seen immediately and most positively from the subsequent evacuation of excrement. "In a word, these infusoria gobble sugar, and discharge ethyl alcohol from the intestine and carbon dioxide from the urinary organs." This attitude of total rejection on the part of the foremost chemists is understandable. It was indeed only a few years earlier (1828) that Wöhler had succeeded in artificially producing urea, a substance which had previ- ously been conceived as a type of all substances produced in animal bodies Justus Liebig (1803-1873). Unrivalled pioneer in the field of organic chemistry, who by his studies on plant nutrition first provided a rational basis for agriculture; the inspiring teacher, to whom Germany owes her first teaching laboratory. only, under the influence of the life force. No sooner had it been realized, said Liebig, that all life processes in plants, just as in animals, must be con- ceived as physical and chemical processes, than along came unscientific people and tried to make acts of life out of simple chemical processes. Attempts were made to produce an explanation of a purely chemical na- ture. Berzelius assumed that yeast caused the decomposition of sugar cataly- tically, simply by its presence as a contact substance or catalyst. There seemed to be analogies with many processes-for instance, with the action of very finely divided platinum on hydrogen peroxide which, in the presence of that contact substance, rapidly decomposes into water and oxygen, whilst the platinum apparently remains unchanged. Liebig gave as his opinion that yeast caused fermentation "in consequence of a progressive disintegration 106 1907 E. BUCHNER which it suffers in the presence of air in contact with water" 3. A disintegra- ting body would possess the ability to cause the same change in another substance touching it 4. In their views, therefore, the only part played by the yeast was that of an organic compound in a state of continuous decomposi- tion. The idea of their nature being that of living plants, however, was unilat- erally ignored by the chemical authorities of the day. The experimental investigations of the next decade, by E. Mitscherlich, H. Helmholtz, H. Schröder and others, even though they favoured the vital- Louis Pasteur (1822-1895). Brilliant chemist and bacteriologist, who at the age of 22 investigated the isomerism of racemic and tartaric acid and hereby laid the foundation for a concept of the structure of molecules; who recognized micro-organisms not only as instigators of fermentation phenomena but also of infectious diseases; who by his discovery of vaccination against rabies has served all humanity. istic concept, did not produce complete clarity. More extensive and more convincing indeed were the investigations of T. Bail in Danzig which, how- ever, had unfortunately to be broken off for external reasons before any final conclusion had been reached. It was only the systematic and striking experiments of Louis Pasteur, ex- tending over a decade, which finally led to the recognition that in Nature without living organisms, without live yeast, no fermentation exists. This put an end to all disputes. Fermentation was seen to be a physiological act inseparably linked with the life processes of yeast. Endeavours were now made to comprehend the phenomenon of fermen- CELL-FREE FERMENTATION 107 tation biologically and establish its origin in greater detail. Schwann had already conjectured that sugar fermentation coincided with the feeding pro- cesses of the yeast. A simpler assumption was made by Moritz Traube in Berlin 5 (1858), according to which there was in micro-organisms a certain chemical body which caused fermentation. Similar substances, chemically very active, which are known today as enzymes, had already been traced on many occasions in vegetable and animal bodies - for instance, diastase in ger- minating barley, which converts starch into sugar; pepsin or peptase, found Moritz Traube (1826-1894). Received comprehensive academic training, but was forced by circumstances to become a wine-merchant for many years. He then became active as a private scientist in Berlin: science owes to him a large number of valuable investigations, in particular on oxidation phenomena and on biological problems. in gastric juice by T. Schwann, which converts into solution and digests co- agulated protein; and invertase, discovered by Berthelot in yeast cells, which decomposes cane sugar into glucose and fructose. This enzyme theory as an explanation of fermentation phenomena found great favour in wide circles. Berthelot, Claude Bernard, Schönbein and Schaer, F. Hoppe-Seyler, G. Hüfner and particularly also Liebig supported it. The plant physiologists, however, principally Naegeli and Sachs, raised weighty objections. In particular, every attempt made to separate such an enzyme from the yeast cells had failed. In spite of extensive investigations, M. Berthelot, Adolf Mayer, Naegeli and Low, even Pasteur himself, the great experimenter had no success. Pasteur’s words sound extremely resign- ed: 6 "In what does for me the chemical process of sugar decomposition con- 108 1907 E.BUCHNER sist, and what is its intrinsic cause? I confess that I am completely in the dark about it. Can we say that the yeast nourishes itself on the sugar, only to give it off again as an excrement in the form of alcohol and carbon dioxide? Or must we say that the yeast in its development produces a substance of the nature of peptase which acts on the sugar and disappears as soon as it has ex- hausted itself, since we find no substance of this kind in the fermentation liquids ? I have no answer to the substance of these hypotheses.